DETERMINANTS OF APICAL MEMBRANE FORMATION AND DISTRIBUTION IN MULTICELLULAR EPITHELIAL MDCK CYSTS

Madin-Darby canine kidney epithelial cells form three-dimensional cyst s in spinner culture with a defined cell surface polarity. Transfer of cysts from spinner culture to a collagen gel matrix results in rapid loss of apical membrane proteins from the outside surface of the cyst, degradation of extracellular matrix (ECM) from the cyst lumen, and de novo formation of the apical membrane at the luminal surface. Degrada tion of endogenous ECM was inhibited with 1,10-phenanthroline, an inhi bitor of metalloproteinases, resulting in cysts in which cells are sur rounded by either cell-cell or cell-substratum contacts. The consequen ce of the lack of a free cell surface on the formation of a new apical membrane domain in these cysts was analyzed. Changes in cell surface polarity were followed with antibodies to marker proteins of the apica l or basolateral membranes. In the absence of a free cell surface, the apical membrane formed de novo by accumulation and fusion of presorte d vesicles containing apical membrane proteins; the coalescence of the se vesicles results in the formation of a central lumen. These results provide novel insights into the generation of membrane domains and fo rmation of a lumen in complex, three-dimensional epithelial structures in development.